Gene therapy is the transfer of genetic material into mammalian cells to treat disease. Numerous diseases such as lysosomal storage diseases which involve cells of the monocytic lineage could be ammenable to gene therapy. Pluripotent stem cells are attractive targets for gene therapy in such diseases. Transplantation of engineered stem cells would result in the continuous presence of the gene in all blood cells for the life of the organism. However, there are many limitations to gene therapy in hematopoietic stem cells. One restriction to the use of retroviral vectors for gene transfer is that cell division is required for proviral integration into the target cell. Thus genes can not be maintained in nonreplicating cells; such as quiescent stem cells. A second limitation is the continued high level expression of the transferred gene. Monocytic precursors that are not quiescent should be a more desirable target cell population for gene therapy than the pluripotent stem cell. Once infected, these cells could be transplanted into recipient animals where they would take up residence in liver or spleen (or other tissues) and differentiate into tissue macrophages expressing the desired gene. A key question is whether early cells committed to the monocytic lineage can serve as progenitors of tissue macrophages and result in long term engraftment at specific tissue sites. The specific aims of this proposal are: 1. To determine the kinetics of tissue macrophage and microglial engraftment following bone marrow transplantation. This will be done using the ROSA 26 transplantation model which can detect individual transplanted cells. 2. To determine if cells other than quiescent hematopoietic stem cells can serve as progenitors of monocytes and tissue macrophages in a mouse transplantation model. These studies will initially be done with the ROSA 26 transplantation model. Later studies will involve the use of retroviral vectors as a model of gene therapy. 3. To extend these studies to a preclinical large animal model (eat) using retrovirally marked cells in both postnatal and in utero transplantation systems.